PROJECT SUMMARY/ABSTRACT Mucormycosis, most commonly caused by Rhizopus oryzae, is a lethal infection in patients with increased available serum iron (e.g. from diabetes), or in patients with neutropenia or steroid-treatment. The population- based incidence rose by more than 50% during the first decade of the 21st century. Unfortunately, the mortality rate of mucormycosis has remained ?40%. New strategies to combat mucormycosis are urgently needed. Using small molecule iron chelators, we found that iron sequestration can protect diabetic mice from lethal mucormycosis. However, animal models showed that small molecule chelation therapy was much less effective, and had substantial toxicity, in neutropenic mice. Consistent with this finding, the follow on clinical trial was enriched for neutropenic patients with mucormycosis, and had very few diabetics--the drug failed. The current application is based on a novel biological approach. Numerous studies have demonstrated the ability of transferrin to restrict microbial growth via iron sequestration. We showed that treatment of mice infected with Staphylococcus aureus, Acinetobacter baumannii, and Candida albicans markedly improved survival. A major advantage of transferrin over small molecules is its safety. Small molecule chelation alters iron excretion patterns, chelating iron away from myeloid cells, which are normally the predominant recyclers of iron in the host, with resulting hematopoietic toxicity, and delivering iron to renal tubules where iron does not normally get excreted, with resulting nephrotoxicity. Thus, serious toxicity to bone marrow, kidneys, and other organ systems can occur, which offsets efficacy benefit in neutropenic hosts. In contrast, administration of transferrin utilizes and enhances normal myeloid iron sequestering routes in the body, and thus may overcome limitations of small molecule chelation, avoiding myeloid and nephrotoxicity in such patients. Specific Aims: 1. To dose-optimize rhTransferrin in treatment of mucormycosis in diabetic, steroid-treated, or neutropenic mice. Two rounds of transferrin dose optimization will be studied vs. placebo. The primary efficacy endpoint will be time to moribund condition, and secondary endpoints will include tissue fungal burden, inflammatory cytokines, and tissue histopathology with rhTransferrin vs. placebo. 2. To determine the impact of combination therapy with rhTransferrin plus liposomal amphotericin with or without an echinocandin during mucormycosis in diabetic, steroid-treated, or neutropenic mice. The efficacy of rhTransferrin alone vs. plus liposomal amphotericin (LAmB) vs. plus both LAmB and an echinocandin, will be compared to LAmB plus echinocandin without transferrin (positive control). Outcomes will include time to moribund condition, fungal burden, inflammation, & histopathology. The results of these experiments will support a follow-on R01 to define mechanism of protection, breadth of protection across fungal species and genera, and pharmacodynamics. Furthermore, results will support rapid translation of transferrin as an adjunctive therapy for mucormycosis in patients.